This project will investigate structure-activity relations in the human glycoprotein hormones from two structural standpoints: primary amino acid sequence and post-translational modifications. To define the role of subunit sequence regions in target-organ action (Aim 1) we will: (a) prepare chemically modified subunits of human choriogonadotropin (hCG) for testing of recombination, receptor binding, adenylate cyclase activation and steroidogenesis. The results are expected to provide information on the importance of several amino acid sequence positions unique to the human hormones. (b) Synthesize several peptide fragments for use in preparing sequence-specific antisera for neutralization studies and evaluation of conformational changes. Two peptides with potential for amphiphilic helix formation will also be tested for direct interaction with ovarian membrane receptors. A post-translational modification clearly important in the action of hCG is the asparagine-linked side-chain complex carbohydrate. Extending our previous work characterizing the biological effects of deglycosylated hCG, we will explore the biochemical basis for the interaction of carbohydrate with cell membrane components to influence "coupling" of receptor to postreceptor events (Aim 2). We will determine requirements for carbohydrate specificity by preparing and testing hCG with modified, otherwise intact oligosaccharides, and will attempt to remove the chains intact from the subunit for assay of binding and cyclase activity. We will use carbohydrate coupling procedures to add oligosaccharide chains to specific locations on deglycosylated or intact hCG subunits. These will be tested for their ability to recombine and elicit hCG responses in ovarian cells. If bioactive products are obtained, we will vary the configuration of added carbohydrate for define specificity. Finally, evidence for a subunit conformational change after deglycosylation will be studied by use of the region-specific antisera as probes for alterations involving different regions of the molecule. Through understanding the way in which these structural features mediate hormone action we hope to suggest new directions for design of antagonists, or potent agonists, for a range of potential clinical applications in reproductive endocrinology.